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Noggin BMP-7 Protein Complex
Hannah Regan '12


I. Introduction

In embryonic development, one of the most basic and important processes is axis patterning, that is, which side of the organism will be its front, its two sides, its top and bottom. Within the organism, axis patterning determines the structure of tissues as well as their placement. This patterning is often achieved through protein gradients (such as the bicoid gradient4 in Drosophila ) of signaling ligands. On one end of the axis there will be a high protein concentration that fades until it is essentially non-existent at the other end of the axis. The concentration of the protein sends signals of varying strength to cells that cause them to follow a certain developmental pathway.

This gradient is often achieved through antagonism of the signal protein by another ligand: the signal protein decreases in expression along the axis because it is repressed by a high concentration of a protein that binds the signal and inhibits its function.

Bone Morphogenetic Proteins (BMPs) are extracellular signaling proteins that play crucial roles in both embryonic development and adult tissue regeneration processes. One of the evolutionarily oldest signaling pathways, BMPs are required for patterning of the embryonic axes. When a BPM ligand binds to a surface receptor protein on a cell, it initiates a signal cascade (such as the SMAD pathway6 ), with the end result of switching on or off certain genes in the nuclear DNA. BMPs are regulated by various classes of antagonists that can block its binding interface and prevent it from contacting the cell surface receptors.

Noggin is believed to have evolved from the same ancestral protein as BMP72 . It was implicated as an important developmental protein when its ability to rescue Xenopus embryos that had been ventralized was discovered1. It is an extracellular cystine- knot protein that exclusively binds BMP and thus inhibits its function6. Noggin is required for proper neural tissue development in embryos and proper differentiation of the embryonic axes.

II. BMP-7 Structure

BMP-7 is a homodimer with two wing shaped monomers, each of which is anchored in a cystine-knot core. Each 'wing' is formed by two pairs of antiparallel ß-strands that are referred to as finger 1 and finger 2. There is one α-helix per monomer that extends for 3.5 turns . 3

The conformation of each monomer is stabilized in two main ways: through contacts between the fingers, and by the cystine-knot conformation. Finger 1 contains a 13 residue Ω-loop with five polar residues that are in contact with the solvent and six non-polar residues that contribute, along with eight residues (also non-polar) from finger 2, to the stability of the interface between the fingers 3.

Each wing is anchored in a 'core' region that contains a ten-membered cystine knot that is composed of eight residues linked by two disulfide bonds between cystine residues . A third disulfide bond passes through the eight membered ring to form the ten membered knot 2.

The BMP-7 dimer is formed by the interactions between the α-helix of each monomer and the concave shape formed by the finger region of the opposite monomer and by an interchain disulfide bond between cystine residues 103 on each chain .

Upon binding by Noggin, BMP-7 undergoes a conformational change, each wing goes from relatively flat conformation to a curled conformation

III. Noggin Structure

Similar to BMP-7, Noggin is a dimer, the C-terminal domain of each monomer has two regions of β-sheets that are refered to as finger 1 and finger 2. The N-terminal region of the monomer is about 20 residues long, and is referred to as the clip domain, a helical domain extends from finger 1 up to the part of the monomer that participates in dimerization 2.

The cystine knot in Noggin also serves to stabilize the conformation of the protein and maintain its integrity. In Noggin, unlike BMP, the cystine knot is made up of a ten-membered ring that has two disulfide bonds . A third disulfide bond between two cystine residues not in the ring structure passes through it to complete the knot residues that are not in the ring structure.

The noggin dimer is formed by contact between α-helix 4 of each monomer and by the disulphide bond between the cystine residue 232 of each monomer .

Noggin has a heparan binding domain between α-helix 3 and α-helix 4 . This domain binds to heparan sulfate proteoglycans 5on cell surfaces, which may play a key role in determining the Noggin gradient in the extracellular matrix2.

IV. Noggin BMP-7 Antagonism

BMP-7 has a hydrophobic 'pocket' of 5 residues where it binds to receptors . The clip of the Noggin antagonist has a proline residue, the hydrophobic ring of which fits into this pocket and anchors Noggin to BMP-7 as well as prevents BMP-7 from binding its intended receptors 2.

BMP-7 has another receptor binding region defined by 5 residues . Blocked by the C-terminal end of the clip domain and by the tips of fingers 1 and 2 2.

V. Noggin Mutations

There are three disease causing mutations that are identified in Noggin that result in apical joint fusion. Proximal symphalangism is the result of six residue point mutations: Pro35Arg, Cys184Tyr, Gly189Cys, Ile220Asn, Tyr222Cys/Asn, Pro233Leu . These mutations likely interfere with Noggin's ability to form a dimer. Tarsal/carpal coalition syndrome is the result of three point mutations: Arg204Leu, Pro35Arg, Tyr222Cys . These mutations prevent Noggin from binding effectively to BMP-7. Multiple syntosis syndrome is the result of one point mutation: Trp217Gly

Without proper function, Noggin cannot bind BMP-7 to inhibit its function and establish a protein gradient. This results in mutations in bone development in embryos.

VI. References

[1] De Robertis, E.M., Kuronda, H. 2004. Dorsal- Ventral Patterning and Neural Induction in Xenopus Embryos. Annu. Rev. Cel Dev. Biol. 20: 258-308.

[2] Groppe, J., Greenwald, J. Wiater, E., Rodriguez-Leon, J., Economides, A.N., Kwiatkowski, W., Affolter, M., Vale, W.W., Izpisua Belmonte, J.C., Choe, S. 2002. Structural basis of BMP signalling inhibition by the cystine knot protein Noggin. Nature 420: 636-648.

[3] Griffith, D.L., Keck, P.C., Sampath, T.K., Rueger, D.C., Carlson, W.D. 1996. Three-dimensional structure of recombinant human osteogenic protein 1: Structural paradigm for the transforming growth factor Β Superfamily. Proc. Natl. Acad. Sci. USA 93: 878-883.

[4] Lipshitz, H.D. 2009. The diffusion-based model for formation of the Bicoid protein gradient. Diagram. Nature Reviews 10: 509-512[509].

[5] Paine-Saunders, S., Viviano, B.L., Economides, A.N., Saunders, S. 2002. Heparan sulfate proteoglycans retain Noggin at the cell surface: a potential mechanism for shaping bone morphogenetic protein gradients. Journal of Biological Chemistry 277: 2089-2096.

[6] Walsh, D.W., Godson, C., Brazil, D.P., Martin, F. 2010. Extracellular BMP-antagonist regulation in development and disease: tied up in knots. Trends in Cell Biology 20: 2440256.

Silva, Nathan and David Marcey. Intro to Jmol Scripting.

Acknowledgements to Karen Hicks' Developmental Biology class for background information on developemental processess outlined in my introduction.

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